Mechanism of cytotoxicity of ricin

蓖麻毒素的细胞毒性机制

• 批准号: 7630083
• 负责人: NILGUN E TUMER
• 金额: $ 3.64万
• 依托单位: RUTGERS, THE STATE UNIV OF N.J.
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-03-15 至 2012-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7630083
• 关键词: Address; Adenine; Aerosols; Antidotes; Apoptosis; Apoptotic; Binding; Biochemical; Biological Models; Biological Warfare; Bioterrorism; Breathing; Categories; Cell Death; Cells; Cessation of life; Cytosol; Defect; Depurination; Development; Endocytosis; Endoplasmic Reticulum; Enterotoxins; Genes; Genome; Genomics; Induction of Apoptosis; Intoxication; Lectin; Lung; MAPK14 gene; MAPK8 gene; Mammalian Cell; Measures; Medical; Membrane; Messenger RNA; Molecular; Mutagenesis; N glycosidase; National Institute of Allergy and Infectious Disease; Plants; Plasmids; Process; Protein Biosynthesis; Protein Synthesis Inhibition; Proteins; RNA Splicing; Resistance; Respiratory System; Ribosomal RNA; Ribosomes; Ricin; Ricinus communis; Risk; Role; Signal Pathway; Signal Transduction; Small Interfering RNA; System; Techniques; Terrorism; Toxin; Translations; Vaccines; Yeasts; base; biodefense; cell injury; cell killing; cytotoxicity; design; instrument; mutant; novel therapeutics; prevent; prototype; response; ribosomal A-protein

Ribosome inactivating proteins (RIPs) have been used as instruments of biological warfare and terrorism. Ricin is a heterodimeric plant toxin that consists of A and B-chains and the prototype of type II RIPs. Its B- chain is a lectin that enables cell binding. After endocytosis, the A-chain translocates through the endoplasmic reticulum membrane to reach the cytosol where its N-glycosidase activity removes a specific adenine from the highly conserved, sarcin/ricin loop (SRL) in the large rRNA. Ricin has been classified as Category B priority for biodefense. Very little is known about the membrane translocation and ribosome interactions of ricin and the molecular mechanism by which it causes apoptosis in mammalian cells. We have established yeast as a biologically relevant model system to study the activity of RTA, and isolated mutant forms of RTA, which do not kill cells. Translation inhibition by ribosome depurination has been hypothesized to be responsible for the cytotoxicity of ricin. However, our preliminary analysis of the nontoxic RTA mutants indicates that ribosome depurination is necessary, but not sufficient for cytotoxicity. We further showed that wild type RTA inhibits induction of the unfolded protein response (UPR) in yeast and in mammalian cells and induces activation of JNK, p38 and ERK signaling pathways in mammalian cells. This project aims to use yeast and mammalian cells as complementary systems to understand the molecular basis for ricin intoxication. Specific Aims: 1. Using the nontoxic RTA mutants, determine if ribosome binding and depurination are required for cytotoxicity in yeast and induction of apoptosis in mammalian cells. 2. Characterize the interaction between RTA and ribosomal protein POand determine if binding to POis essential for ribosome depurination by RTA. 3. Determine if RTA causes cell death by inhibiting the development of UPR in yeast and in mammalian cells. 4. Identify the cellular genes necessary for cytotoxicity of RTA in yeast. Ricin is not only a bioterrorism threat, but inhibits translation by a similar mechanism as the bacterial enterotoxins. Therefore, the studies outlined in this application will have important implications for the design of protection strategies against AB-toxins that are classified as high-risk candidates for bioterrorism.

核糖体失活蛋白(RIPs)已被用作生物战和恐怖主义的工具。 蓖麻毒素是一种由A链和B链组成的异二聚体植物毒素，是II型RIPs的原型。它的B- 链是一种使细胞结合的凝集素。在内吞作用后，A链通过 内质网膜到达胞浆，在那里其N-糖苷酶活性去除特定的 腺嘌呤来自于大rRNA中高度保守的sarcin/ricin环(SRL)。蓖麻毒素被归类为 生物防御的B类优先事项。对膜转位和核糖体知之甚少。 蓖麻毒素的相互作用及其导致哺乳动物细胞凋亡的分子机制。我们 建立了酵母作为与生物相关的模型系统来研究RTA的活性，并分离出 RTA的突变形式，它不会杀死细胞。核糖体脱嘌呤对翻译的抑制作用 推测与蓖麻毒素的细胞毒性有关。然而，我们初步分析了无毒的 RTA突变体表明，核糖体脱嘌呤是必要的，但不足以产生细胞毒性。我们进一步 结果表明，野生型Rta抑制酵母和酵母中未折叠蛋白反应(UPR)的诱导 并诱导哺乳动物细胞中JNK、p38和ERK信号通路的激活。这 该项目旨在利用酵母和哺乳动物细胞作为互补系统来理解分子 蓖麻毒素中毒的基础。具体目的：1.利用无毒的RTA突变体，确定核糖体是否 结合和去嘌呤是酵母细胞毒性和诱导哺乳动物细胞凋亡所必需的。 2.鉴定RTA与核糖体蛋白POI的相互作用，并确定是否与POI结合 RTA对核糖体去嘌呤是必需的。3.确定RTA是否通过抑制细胞死亡而导致细胞死亡 UPR在酵母和哺乳动物细胞中的发展。4.确定细胞所需基因 RTA在酵母中的细胞毒性。蓖麻毒素不仅是生物恐怖主义的威胁，而且还能抑制类似的 其作用机制与细菌肠毒素有关。因此，本申请书中概述的研究将具有 对高危AB类毒素防护策略设计的重要启示 生物恐怖主义的候选人。